1. Field of the Invention
The present invention generally relates to polypeptides comprising an antibody sequence linked to a cytokine sequence and methods of making and using thereof.
2. Description of the Related Art
Interleukin 12 (IL-12) is a heterodimeric cytokine composed of p35 and p40 subunits which are encoded by 2 separate genes, IL-12A and IL-12B, respectively. The natural activity of IL-12 is likely influenced by its heparin binding activity which retains IL-12 close to local tissue compartments to execute its paracrine effects. See e.g. Hasan et al. (1999) J. Immunol. 162:10064-1070.
Both IL-2 and IL-12 have been used for the immunotherapy of cancer. Antibody-(IL-2) fusion proteins have been used in successful anti-tumor experiments using animal models. See Penichet & Morrison (2001) J Immunol Met 248:91-101. Numerous studies have explored various combinations of antibodies and, e.g., IL-2, as direct targeting agents of tumor cells. For example, a tumor specific antibody-(IL-2) fusion protein was previously developed, and comprised a human IgG3 specific for the idiotype (Id) of the immunoglobulin expressed on the surface of the B cell lymphoma 38C13 with human IL-2 fused at the end of the CH3 domain. See Penichet et al. (1998) J Interferon Cytokine Res 18:597-607. Other examples of antibody fusion molecules include a humanized anti-HER2/neu IgG3 fused to IL-12 and a recombinant IgG3-(IL-2) fusion protein. See Peng et al. (1999) J Immunol 163:250-8); and Penichet et al. (2001) Human Antibodies 10:43-49.
In all of the above work, it is important to note that the antibody-cytokine fusion proteins containing IL-2 and IL-12 have been used as direct anti-tumor agents which are expected to directly targeted tumors in animal models. The antibody fusion proteins bind to antigens on tumor surfaces, thus increasing the local concentration of the antibody-cytokine fusion protein in the tumor microenvironment.
Unfortunately, although anti-tumor responses were observed in some subjects, the systemic injection of free IL-2 or IL-12 often results in severe toxicity due to the high dose that is required to induce the anti-tumor immune response. Although fusing the cytokine to an antibody is expected to better target the cytokine into the tumor reducing its site effects (toxicity) we have also found that these antibody-cytokine fusion proteins also exhibit the heparin-binding activity of the fused cytokines (IL-2 or IL-12), which makes them bind non-specifically to a broad variety of cells throughout the body and the extracellular matrix (ECM) of normal tissues. This heparin-binding activity is expected to reduce the capacity of the antibody-cytokine fusion proteins to specifically target the tumors due to its retention outside the tumor microenvironment, which may also result in unfavorable side effects.
Therefore, a need still exists for effective therapeutics.